Deleterious Effect of the Q o Inhibitor Compound Resistance-Conferring Mutation G143A in the Intron-Containing Cytochrome b Gene and Mechanisms for Bypassing It

Abstract

The mutation G143A in the inhibitor binding site of cytochrome b confers a high level of resistance to fungicides targeting the bc ₁ complex. The mutation, reported in many plant-pathogenic fungi, has not evolved in fungi that harbor an intron immediately after the codon for G143 in the cytochrome b gene, intron bi2. Using Saccharomyces cerevisiae as a model organism, we show here that a codon change from GGT to GCT, which replaces glycine 143 with alanine, hinders the splicing of bi2 by altering the exon/intron structure needed for efficient intron excision. This lowers the levels of cytochrome b and respiratory growth. We then investigated possible bypass mechanisms that would restore the respiratory fitness of a resistant mutant. Secondary mutations in the mitochondrial genome were found, including a point mutation in bi2 restoring the correct exon/intron structure and the deletion of intron bi2. We also found that overexpression of nuclear genes MRS2 and MRS3 , encoding mitochondrial metal ion carriers, partially restores the respiratory growth of the G143A mutant. Interestingly, the MRS3 gene from the plant-pathogenic fungus Botrytis cinerea , overexpressed in an S . cerevisiae G143A mutant, had a similar compensatory effect. These bypass mechanisms identified in yeast could potentially arise in pathogenic fungi.